BrdU Proliferation Assay for MCF-7 and MDA-MB-231 Cells

Cell proliferation was analyzed by the BrdU incorporation ELISA assay following the manufacturing instructions (Roche Diagnostics GmbH, Mannheim, Germany). The incorporation of BrdU into genomic DNA during cell growth was quantified and analyzed after SPGE extract treatment (10–1,000 μg/mL) in our experiments. After the substrate degradation, the color intensity changes were read at 450 nm (reference wavelength: 690 nm) by Cytation^TM 3 Cell Imaging Multi-Mode Reader (Biotec). The numbers of experimental repeats were set (n = 3). Based on Resazurine and BrdU assays, the final IC₅₀ was calculated and determined: 160 μg/mL (MCF-7 cells) or 130 μg/mL (MDA-MB-231 cells) (Kubatka et al., 2019 (link); Kubatka et al., 2020a (link); Kubatka et al., 2020b (link)).